Abstract Title:

Matrix Metalloproteinases Activation by Alpha 2- Macroglobulin in Glial Müller Cells

Presentation Start/End Time:

Sunday, Apr 27, 2008, 2:30 PM - 4:15 PM

Location:

Hall B/C

Reviewing Code:

318 retina: biochemistry and molecular biology (non-photoreceptor) - BI

Author Block:

M.C. Sanchez¹, P.F. Barcelona¹, S.G. Ortiz¹, J.D. Luna², C.P. Juarez², G.A. Chiabrando¹. ¹CIBICI-Dpto de Bioquimica Clinica, Fac de Ciencias Quimicas UNC, Cordoba, Argentina; ²Departamento de Oftalmologia, Fundación VER, Cordoba, Argentina.

Keywords:

602 Muller cells, 659 proteolysis, 711 signal transduction

Purpose: Glial Muller cells (MC) are known to undergo functional and morphological changes and to produce matrix metalloproteinases (MMPs) during retinal proliferative disorders. In previous in vivo and in vitro studies we have demonstrated that MC express the alpha 2- macroglobulin (α₂-M) receptor (LRP-1) and α₂-M induces MMP-2 activity in primary cultures of MC. Herein we investigated the biochemical mechanism of α₂-M effect on MMPs activity regulation, in an spontaneously immortalized human Müller cell line that express LRP-1.
Methods: The human MIO-M1 (a kind gift from G. A. Limb, Institute of Ophthalmology and Moorfields, Eye Hospital, London, UK) was cultured in the absence or presence of 60 nM α₂-M for different periods of time. The effect of α₂-M on MMPs activity was evaluated in aliquots of MC supernatants by zymographic analysis. The expression level of MT1-MMP was analyzed by RT-PCR, Western blot and immunofluorescence analysis. MMP-2 and TIMP-2 expressions were also examined by RT-PCR. To evaluate intracellular signaling pathways induced by α₂-M, MIO-M1 cells were pre-treated for 30 min with 20 µM PD98059, 50 µM SP600125, 25 µM SB203580 or 10 µM LY294002 and then treated with α₂-M for 1h.
Results: α₂-M increased MMP-2 activity whereas MMP-9 activity was not modified. By zymographic analysis was observed that the inhibition of JNK and Akt/PI3K, but not MAPK-ERK1/2, modified the α₂-M-induced MMP-2 activity. In addition, α₂-M also increased MT1-MMP and TIMP-2 mRNA and MT1-MMP protein, which was associated with the MMP-2 activity. Finally, we demonstrated by immunofluorescences that under α₂-M induction MT1-MMP was predominantly localized at the cell surface and in the cellular processes of glial cells.
Conclusions: The present study demonstrates that the α₂-M/LRP-1 system regulates the extracellular proteolytic activity of MMP-2 in MIO-M1 cells by inducing the MT1-MMP and TIMP-2 expression, suggesting that JNK and Akt/PI3K pathways are involved.

Commercial Relationship:

 M.C. Sanchez, None; P.F. Barcelona, None; S.G. Ortiz, None; J.D. Luna, None; C.P. Juarez, None; G.A. Chiabrando, None.

Support:

SECyT, FONCyT and CONICET